41.1.28A
Reagents
AOAC Official Method 996.06
Fat (Total, Saturated, and Unsaturated)
in Foods
Hydrolytic Extraction Gas Chromatographic Method
First Action 1996
Revised 2001
(Applicable to determination of fat in foods.)
Caution: Boron trifluoride may be fatal if inhaled.
See Tables 996.06A–C for the results of the interlaboratory study
supporting acceptance of the method.
A. Principle
Fat and fatty acids are extracted from food by hydrolytic methods
(acidic hydrolysis for most products, alkaline hydrolysis for dairy
products, and combination for cheese). Pyrogallic acid is added to
minimize oxidative degradation of fatty acids during analysis.
Triglyceride, triundecanoin (C11:0), is added as internal standard. Fat
is extracted into ether, then methylated to fatty acid methyl esters
(FAMEs) using BF3 in methanol. FAMEs are quantitatively measured
by capillary gas chromatography (GC) against C11:0 internal standard.
Total fat is calculated as sum of individual fatty acids expressed as
triglyceride equivalents. Saturated and monounsaturated fats are
calculated as sum of respective fatty acids. Monounsaturated fat
includes only cis form.
B. Apparatus
(a) Gas chromatograph.—With hy drogen flame ionization
detector, capillary column, split mode injector, oven temperature
programming sufficient to implement a hold-ramp-hold sequence.
Operating conditions: temperature (°C): injector, 225; detector, 285;
initial temp, 100 (hold 4 min); ramp, 3°C/min; final temp 240; hold
15 min; carrier gas, helium; flow rate, 0.75 mL/min; linear velocity,
18 cm/s; split ratio, 200:1.
(b) Capillary column.—Separating the FAME pair of adjacent
peaks of C18:3 and C20:1 and the FAME trio of adjacent peaks of C22:1,
C20:3, and C20:4 with a resolution of 1.0 or greater. SP2560 100 m ´
0.25 mm with 0.20 mm film is suitable.
(c) Mojonnier flasks.
(d) Stoppers.—Synthetic rubber or cork.
(e) Mojonnier centrifuge basket.
(f) Hengar micro boiling granules.
(g) Baskets.—Aluminum and plastic.
(h) Shaker water bath.—Maintaining 70°–80°C.
(i) Steam bath.—Supporting common glassware.
(j) Water bath.—With nitrogen stream supply, maintaining 40° ±
5°C.
(k) Wrist action shaker.—Designed for Mojonnier centrifuge
baskets.
(l) Mojonnier motor driven centrifuge.—Optional; maintaining
600 rpm.
(m) Gravity convection oven.—Maintaining 100° ± 2°C.
(n) Vortex mixer.
(o) Gas dispersion tubes.—25 mm, porosity “A,” extra coarse,
175 mm.
(p) Three-dram vials.—About 11 mL.
(q) Phenolic closed top caps.—With polyvinyl liner, to fit vials.
(r) Teflon/silicone septa.—To fit vials.
C.
(a) Pyrogallic acid.
(b) Hydrochloric acid.—12 and 8.3M. To make 8.3M HCl, add
250 mL 12M HCl to 110 mL H2O. Mix well. Store at room
temperature (20°–25°C).
(c) Ammonium hydroxide.—58% (w/w).
(d) Diethyl ether.—Purity appropriate for fat extraction.
(e) Petroleum ether.—Anhydrous.
(f) Ethanol.—95% (v/v).
(g) Toluene.—Nanograde.
(h) Chloroform.
(i) Sodium sulfate.—Anhydrous.
(j) Boron trifluoride reagent.—7% BF3 (w/w) in methanol, made
from commercially available 14% BF3 solution. Prepare in the hood.
(k) Diethyl ether–petroleum ether mixture.—1 + 1 (v/v).
(l) Triglyceride internal standard solution.—C11:0-triundecanoin;
5.00 mg/mL in CHCl3. Accurately weigh 2.50 g C11:0-triundecanoin
into 500 mL volumetric flask. Add ca 400 mL CHCl3 and mix until
dissolved. Dilute to volume with CHCl3. Invert flask at least
10 additional times. Triglyceride internal standard solution is
stable up to 1 month when stored in refrigerator (2°–8°C).
( m ) F a t t y a c i d m e t h y l e s t e r s ( FA M E s ) s t a n d a rd
solutions.—(1) Mixed FAMEs standard solution.—Reference
mixture containing series of FAMEs, including C18:1 cis and trans
(available as GLC-85 from Nu Chek Prep, Elysian, MN 56028,
USA, or equivalent). To prepare mixed FAMEs standard solution,
break top of glass vial, open, and carefully transfer contents to
3-dram glass vial. Wash original vial with hexane to ensure
complete transfer and add washings to 3-dram glass vial. Dilute to ca
3 mL with hexane.
(2) C11:0 FAME standard solution.—C11:0-Undecanoic methyl
ester in hexane. Use only in preparation of individual FAME
standard solutions, (3). To prepare C11:0 FAME standard solution,
break top of glass vial open and carefully transfer contents to 50 mL
volumetric flask. Wash original vial with hexane to ensure complete
transfer and add washings to 50 mL volumetric flask. Dilute to
volume with hexane. C11:0 FAME standard solution is stable up to
1 week when stored at 0°C.
(3) Individual FAME standard solutions.—Standard solutions of
each of fol low ing FAMEs: C 4 : 0 -tetranoic methyl es ter,
C 6 : 0 -hexanoic methyl es ter, C 8 : 0 -octanoic methyl es ter,
C 10:0 -decanoic methyl es ter, C 12:0 -dodecanoic methyl es ter,
C13:0-tridecanoic methyl ester, C14:0-tetradecanoic methyl ester,
C14:1-9-tetradecenoic methyl ester, C15:0-pentadecanoic methyl
ester, C15:1-10-pentadecenoic methyl ester, C16:0-hexadecanoic
m e t h y l e s t e r, C 1 6 : 1 - 9 - h e xa d e c e n o i c me t h y l e s t e r,
C17:0-heptadecanoic methyl ester, C17:1-10-heptadecenoic methyl
ester, C18:0-octadecanoic methyl ester, C18:1-9-octadecenoic methyl
e s t e r, C 1 8 : 2 - 9 , 1 2 -o c t a d e c a d i e no i c m e th y l e s t e r,
C18:3-9,12,15-octadecatrienoic methyl ester, C20:0-eicosanoic methyl
ester, C20:1-8-eicosenoic methyl ester, C20:2-11,14-eicosadienoic
methyl ester, C20:3-11,14,17-eicosatrienoic methyl ester, and
C22:0-docosanoic methyl ester. Prepare individual FAME standard
solutions as follows: Break top of glass vial open and carefully
transfer contents to 3-dram glass vial. Wash original vial with hexane
to ensure complete transfer and add washings to 3-dram glass vial.
Add 1.0 mL C11:0 FAME standard solution, (2), dilute to total volume
of ca 3.0 mL with hexane. Individual FAME standard solutions are
stable up to 1 week when stored in refrigerator (2°–8°C).
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Table 996.06A. Interlaboratory study results for determination of total fat in food by hydrolytic extraction—gas chromatography
No. of labs/
a sr sR b c RSDr, % RSDR, %
Product x, % outliers r R HorRat
Wheat-based cereal 1.96 — 0.208 0.260 0.582 0.728 10.6 13.3 3.69
Peanut butter 46.3 10/2 0.86 2.37 2.41 6.64 1.85 5.12 2.28
Fish sticks 11.2 10/2 0.354 0.541 0.991 1.51 3.14 4.80 1.73
Parmesan cheese 26.5 — 0.540 4.17 1.51 11.7 2.04 15.8 6.47
Chocolate cake (baked) 13.3 — 0.929 1.95 2.60 5.46 7.00 14.7 5.43
Fruit snack 3.92 10/1 0.087 0.146 0.244 0.409 2.22 3.74 1.15
Ground beef 21.9 10/1 1.11 1.82 3.11 5.10 5.06 8.32 3.31
Yogurt 1.46 — 0.131 0.222 0.367 0.622 8.98 15.2 4.03
a
Blind duplicates.
b
r = 2.8 ´ sr.
c
R = 2.8 ´ sR.

D. Extraction of Fat
Finely grind and homogenize test samples prior to extraction of fat.
[Note: With matrixes of unknown composition, it may be
necessary to analyze test portion without addition of internal
standard to ensure against interferences. Should interfering peak be
found, the area of C11 internal standard peak must be corrected
before performing calculations. Use 2.0 mL chloroform instead of
internal standard solution.]
(a) Foods excluding dairy products and cheese.—Accurately
weigh ground and ho mogenized test portion (con tain ing ca
100–200 mg fat) into labeled Mojonnier flask. Force material into
flask as far as possible. Add ca 100 mg pyrogallic acid, C(a), and
2.00 mL triglyceride internal standard solution, C(l). Add a few
boiling granules to flask. Add 2.0 mL ethanol and mix well until
entire test portion is in solution. Add 10.0 mL 8.3M HCl and mix
well. Place flask into basket in shaking water bath at 70°–80°C set at
moderate agitation speed. Maintain 40 min. Mix contents of flask on
Vortex mixer every 10 min to incorporate particulates adhering to
sides of flask into solution. After digestion, remove flask from bath
and allow to cool to room temperature (20°–25°C). Add enough
ethanol to fill bottom reservoir of flask and mix gently.
(b) Dairy prod ucts.—Ac cu rately weigh ground and
homogenized test portion (containing ca 100–200 mg fat) into
labeled Mojonnier flask. Force material into flask as far as possible.
Add ca 100 mg pyrogallic acid, C(a), and 2.00 mL triglyceride
internal standard solution, C(l). Add a few boiling granules to flask.
Add 2.0 mL ethanol and mix well until entire test portion is in
solution. Add 4.0 mL H2O and mix well. Add 2.0 mL NH4OH, C(c),
and mix well. Place flask into basket in shaking water bath at
70°–80°C set at moderate agitation speed. Maintain 10 min. Mix
contents of flask on Vortex mixer every 5 min to incorporate
particulates adhering to sides of flask into solution. After digestion,
re move flask from wa ter bath and add a few drops of
phenolphthalein. Keep solution basic (pink) with addition of
ammonium hydroxide. Add enough ethanol to fill bottom reservoir
of flask and mix gently.
(c) Cheese.—Accurately weigh ground and homogenized test
portion (containing ca 100–200 mg fat) into labeled Mojonnier
flask. Force material into flask as far as possible. Add ca 100 mg
pyrogallic acid, C(a), and 2.00 mL triglyceride internal standard
solution, C(l). Add a few boiling granules to flask. Add 2.0 mL
ethanol and mix well until entire test portion is in solution. Add
4.0 mL H2O and mix well. Add 2.0 mL NH4OH, C(c), and mix well.

Table 996.06B. Interlaboratory study results for determination of saturated fat in food stuffs by hydrolytic extraction—
gas chromatography
No. of labs/
a sr sR b c RSDr, % RSDR, %
Product x, % outliers r R HorRat
Wheat-based cereal 0.493 10/0 0.0391 0.0522 0.109 0.146 7.92 10.6 2.39
Peanut butter 8.72 10/1 0.257 1.81 0.720 5.07 2.95 20.7 7.18
Fish sticks 3.00 10/1 0.223 0.572 0.624 1.60 7.44 19.1 5.64
Parmesan cheese 17.4 — 0.311 2.46 0.871 6.89 1.79 14.1 5.42
Chocolate cake (baked) 3.56 — 0.171 0.304 0.479 0.851 4.81 8.55 2.59
Fruit snack 1.27 10/2 0.0242 0.0362 0.0678 0.101 1.90 2.83 0.74
Ground beef 9.98 10/1 0.636 1.39 1.78 3.89 6.38 13.9 4.92
Yogurt 0.986 — 0.119 0.170 0.333 0.476 12.1 17.2 4.30
a
Blind duplicates.
b
r = 2.8 ´ sr.
c
R = 2.8 ´ sR.

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Table 996.06C. Interlaboratory study results for determination of monounsaturated fat in food stuffs by hydrolytic extraction—
gas chromatography
No. of labs/
a sr sR b c RSDr, % RSDR, %
Product x, % outliers r R HorRat
Wheat-based cereal 0.280 — 0.0320 0.0560 0.0896 0.157 11.4 20.0 4.14
Peanut butter 22.3 10/2 0.411 1.11 1.15 3.11 1.84 4.96 1.98
Fish sticks 1.83 — 0.165 0.313 0.462 0.876 9.02 17.1 4.69
Parmesan cheese 6.43 10/1 0.271 1.09 0.759 3.05 4.20 17.0 5.63
Chocolate cake (baked) 3.79 — 0.413 1.27 1.16 3.56 10.9 33.5 10.25
Fruit snack 1.08 10/2 0.0453 0.734 0.127 2.06 4.17 67.7 17.16
Ground beef 8.88 — 0.930 1.96 2.60 5.49 10.5 22.0 7.65
Yogurt 0.345 10/1 0.0222 0.0542 0.0621 0.152 6.42 15.1 3.23
a
Blind duplicates.
b
r = 2.8 ´ sr.
c
R = 2.8 ´ sR.

Place flask into basket in shaking water bath at 70°–80°C set at
moderate agitation speed. Maintain 20 min. Mix contents of flask on
Vortex mixer every 10 min to incorporate particulates adhering to
sides of flask into solution. Add 10.0 mL 12M HCl and place flask
into boiling steam bath and maintain 20 min. Mix flask contents
every 10 min using Vortex mixer. Remove flask from steam bath and
allow to cool to room temperature (20°–25°C). Add enough ethanol
to flask to fill bottom reservoir and mix gently.
(d) Extraction.—Add 25 mL diethyl ether to Mojonnier flask
from (a), (b), or (c). Stopper flask and place in centrifuge basket.
Place basket in wrist action shaker, securing flask in shaker with
rubber tubing. Shake flask 5 min. Rinse stopper into flask with
di ethyl ether–pe tro leum ether mix ture, C(k). Add 25 mL
petroleum ether, stopper flask, and shake 5 min. Centrifuge flask
(in basket) 5 min at 600 rpm. (Note: If centrifuge is not available,
allow contents to set at least 1 h until upper layer is clear.) Rinse
stopper into flask with diethyl ether–petroleum ether mixture.
Decant ether (top) layer into 150 mL beaker and carefully rinse lip
of flask into beaker with diethyl ether–petroleum ether mixture.
Slowly evaporate ether on steam bath, using nitrogen stream to aid
in evaporation. Residue remaining in beaker contains extracted fat.
E. Methylation
Dissolve extracted fat residue in 2–3 mL chloroform and 2–3 mL
diethyl ether. Transfer mixture to 3 dram glass vial and then
evaporate to dryness in 40°C water bath under nitrogen stream. Add
2.0 mL 7% BF3 reagent, C(j), and 1.0 mL toluene, C(g). Seal vial
with screwcap top containing Teflon/silicone septum. Heat vial in
oven 45 min at 100°C. Gently shake vial ca every 10 min. (Note:
Evaporation of liquid from vials indicates inadequate seals; if this
occurs, discard solution and repeat the entire procedure.) Allow vial
to cool to room temperature (20°–25°C). Add 5.0 mL H2O, 1.0 mL
hexane, and ca 1.0 g Na2SO4, C(i). Cap vial and shake 1 min. Allow
layers to separate and then carefully transfer top layer to another vial
containing ca 1.0 g Na2SO4. (Note: Top layer contains FAMEs
including FAME of triglyceride internal standard solution.)
Inject FAMEs onto GC column or transfer to autosampler vial for
GC analysis.
F. GC Determination
Relative retention times (vs FAME of triglyceride internal
standard solution) and response factors of individual FAMEs can be
obtained by GC analysis of individual FAME standard solutions and
mixed FAME standard solution. Inject ca 2 mL each of individual
FAME standard solutions and 2 mL of mixed FAMEs standard
so lu tion. Use mixed FAMEs stan dard so lu tion to op ti mize
chromatographic response before injecting any test solutions. After
all chromatographic conditions have been optimized, inject test
solutions from E.
G. Calculations
Total fat is the sum of fatty acids from all sources, expressed as
triglycerides. Expressing measured fatty acids as triglycerides
requires mathematical equivalent of condensing each fatty acid
with glycerol. For every 3 fatty acid molecules, 1 glyc erol
(HOCH2CHOHCH2OH) is required. Essentially, 2 methylene
groups and 1 methine group are added to every 3 fatty acids.
Calculate retention times for each FAME in individual FAMEs
standard solutions, C(m)(3), by subtracting retention time of C11:0
peak from retention time of fatty acid peak. Use these retention
times to identify FAMEs in mixed FAMEs standard solution. Use
ad di tional FAME solu tions (from the same sup plier) when
necessary for complete FAME identity verification.
(a) Calculate response factor (Ri) for each fatty acid i as follows:
Psi W
Ri = ´ C11:0
PsC11:0 Wi
where Psi = peak area of individual fatty acid in mixed FAMEs
standard solution; PsC 11:0 = peak area of C11:0 fatty acid in mixed
FAMEs standard solution; WC11:0 = weight of internal standard in
mixed FAMEs standard solution; and Wi = weight of individual
FAME in mixed FAMEs standard solution.
(Note: Peaks of known identity with known relative retention
times are listed in Table 996.06D. When peaks of unknown identity
are observed during the chromatographic run, attempt to identify
such peaks using MS, FTIR, etc. Peaks of unkown identity should
not be included in the summation when quantifying fat in the test
portion.)

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Table 996.06D. Retention time of fatty acids and methyl ester Table 996.06E. Factors (fTG) for conversion of FAMEs to
(FAME) trygliceride equivalents
Relative retention Fatty acid FAi
a
Tri/FAME (FTgi)
b
times
Fatty acid Retention time, min (to 11:0 int. std.) 4:0 Butyric 0.8627 0.9868
4:0 Butyric 10.49 0.46 6:0 Caproic 0.8923 0.9897
6:0 Caproic 12.36 0.54
8:0 Caprylic 0.9114 0.9915
8:0 Caprylic 15.69 0.68
10:0 Capric 20.39 0.89 10:0 Capric 0.9247 0.9928
11:0 Undecanoic 22.99 1.00 11:0 Undecanoic 0.9300 0.9933
12:0 Lauric 25.58 1.11 12:0 Lauric 0.9346 0.9937
13:0 Tridecanoic 28.15 1.22
13:0 Tridecanoic 0.9386 0.9941
14:0 Myristic 30.65 1.33
14:1 Myristoleic 32.63 1.42 14:0 Myristic 0.9421 0.9945
14:1 trans-Myristelaidic 32.01 1.39 14:1 Tetradecenenoic 0.9417 0.9944
15:0 Pentadecanoic 33.04 1.44 15:0 Pentadecanoic 0.9453 0.9948
15:1 Pentadecenoic 34.98 1.52
15:1 Pentadecenoic 0.9449 0.9947
16:0 Palmitic 35.41 1.54
16:1 trans-Palmitelaidic 36.39 1.58 16:0 Palmitic 0.9481 0.9950
16:1 Palmitoleic 36.88 1.60 16:1 Hexadecenoic 0.9477 0.9950
17:0 Margaric 37.54 1.63 17:0 Margaric 0.9507 0.9953
17:1 Margaroleic 38.92 1.69
17:1 Margaroleic 0.9503 0.9952
18:0 Stearic 39.78 1.73
18:1 trans 6-Petroselenic 40.50 1.76 18:0 Stearic 0.9530 0.9955
18:1 trans-Elaidic 40.61 1.77 18:1 Octadecenoic 0.9527 0.9955
18:1 trans 11-Vaccenic 40.72 1.77 18:2 Octadecdieoic 0.9524 0.9954
18:1 Petroselenic 40.90 1.78
18:3 Linolenic 0.9520 0.9954
18:1 Oleic 40.99 1.78
18:1 Vaccenic 41.18 1.79 18:4 Octadectetraenoic 0.9517 0.9954
18:1 Octadecenoic 41.54 1.81 20:0 Arachidic 0.9570 0.9959
18:2 trans-Linolelaidic 41.69 1.81 20:1 Eicosenic 0.9568 0.9959
18:2 trans 9-Linolelaidic 42.11 1.83
20:2 Eicosadienoic 0.9565 0.9958
18:2 trans 12-Linolelaidic 42.53 1.85
18:2 Linoleic 42.87 1.86 20:3 Eicosatrienoic 0.9562 0.9958
20:0 Arachidic 43.75 1.90 20:4 Arachidonic 0.9560 0.9958
18:3 g-Linolenic 44.25 1.92 20:5 Eicosapentaenoic 0.9557 0.9958
20:1 Eicosenic cis 5 44.42 1.93
21:0 Heneicosanoic 0.9588 0.9961
20:1 Eicosenic trans 11 44.45 1.93
20:1 Eicosenic cis 8 44.67 1.94 22:0 Behenic 0.9604 0.9962
20:1 Eicosenic cis 11 44.82 1.95 22:1 Docosaenoic 0.9602 0.9962
20:1 Eicosenic cis 13 44.99 1.96 22:2 Docosadienoic 0.9600 0.9962
18:3 Linolenic 45.02 1.96
22:3 Docosatrienoic 0.9598 0.9961
18:2 Linoleic—conjugated 45.35 1.97
18:2 Linoleic—conjugated 45.40 1.97 22:4 Docosatetraenoic 0.9595 0.9961
21:0 Heneicosanoic 45.69 1.99 22:5 Docosapentaenoic 0.9593 0.9961
18:2 Linoleic—conjugated 46.18 2.01 22:6 Docosahexaenoic 0.9590 0.9961
18:4 Octadectetraenoic 46.39 2.02
23:0 Tricosanoic 0.9620 0.9964
20:2 Eicosadienoic 46.65 2.03
22:0 Behenic 47.46 2.06 24:0 Lignoceric 0.9963 0.9965
20:3 g-Eicosatrienoic 47.94 2.09 24:1 Nervonic 0.9632 0.9965
a
22:1 Cetoleic 48.27 2.10 FAi is the conversion factor for conversion of FAMEs to corresponding fatty
22:1 Erucic 48.50 2.11 acids.
b
20:3 Eicosatrienoic 48.68 2.12 FTgi is the conversion factor for conversion of FAMEs to triglycerides for
20:4 Arachidonic 48.94 2.13 individual fatty acids.
23:0 Tricosanoic 49.22 2.14
22:2 Docosadienoic 50.17 2.18
24:0 Lignoceric 50.79 2.21
20:5 Eicosapentaenoic 50.96 2.22
(b) Calculate amount of individual (triglycerides) (WTG) in test
24:1 Nervonic 51.92 2.26 portion as follows:
22:3 Docosatrienoic 51.98 2.26
22:4 Docosatetraenoioc 52.28 2.27 Pt i ´ Wt C11:0 ´ 10067
.
WFAMEi =
22:5 Docosapentaenoic 54.75 2.38 Pt C11:0 ´ Ri
22:6 Docosahexaenoic 55.82 2.43
WTGi = WFAMEi ´ fTGi

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where Pti = peak area of fatty acid i in test portion; WtC11:0 = weight
of C11:0 internal standard added to test portion, g; 1.0067 =
conversion of internal standard from triglyceride to FAME; PtC11:0 =
peak area of C11:0 internal standard in test portion; and fTGi =
conversion factor for FAMEs to triglycerides for individual fatty
acids (see Table 996.06E).
(Note: If procedure is followed exactly, WtC11:0 should be
0.010 g.)
(c) Calculate amount of total fat in test portion [sum of all fatty
acids, expressed as triglyceride equivalents (including cis and trans
forms of monounsaturated acids)] as follows:
Total fat, % = ( åWTGi ´ 100)/Wtest portion
where Wtest portion = weight of test portion, g.
(d) Calculate weight of each fatty acid (Wi) as follows:
Wi = WFAMEi ´ fFAi
where fFAi = conversion factors for conversion of FAMEs to their
corresponding fatty acids (see Table 996.06E).
(e) Calculate percent of saturated fat in test portion (w/w;
expressed as saturated fatty acids; sum of C4:0, C6:0, C8:0, etc.) as
follows:
Saturated fat, % = (åsaturated Wi/Wtest portion) ´ 100%
(f) Calculate amount of monounsaturated fat in test sample (w/w;
expressed as sum of only cis form of monounsaturated fatty acids
[C16:1, C17:1, C18:1 cis, C20:1, etc.]) as follows:
Monounsaturated fat, % =
(åmonounsaturated Wi/Wtest portion) ´ 100%
Polyunsaturated fat, % =
(åpolyunsaturated Wi/Wtest portion) ´ 100%
(Note: Test samples containing hydrogenated fat will yield
complicated chromatograms due to large number of isomers
formed during hydrogenation process. One general indication of
hydrogenation is presence of C18:1 trans peak(s). For hydrogenated
fat chromatograms, use the following guidelines to calculate FAME
peak areas: trans peaks elute prior to cis, therefore, include all peaks
between C18:1 cis and C18:2 cis,cis in calculation of C18:2 peak area.
Often C18:1 trans “peak” consists of broad series of peaks [due to
positional isomers from hydrogenation]; include all of these in C18:1
trans peak area.)
References: J. AOAC Int. 80, 555(1997); 82, 1146(1999).
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